Comparative dataset on growth, development and reproductive performance of copepod (Cyclops sp.) fed with different microalgae diet

The effects of growth and reproduction on the marine Cyclops sp. were investigated using three microalgae as diets. The development period of Cyclops sp. was evaluated at 106 cells/ml in 15ppt salinity to identify the stationary phase. The survival rate of marine Cyclops from nauplius to adult differed according to the microalgal diet. The results showed that the shortest time (14 days) and highest survival (17.6 ± 0.131 %) for Cyclops sp. was achieved with those fed with Nannochloropsis sp. Whereas, it took longest time (37 days) and lowest survival rate (6.40 ± 0.035 %) when fed Chlamydomonas sp. The developmental period from naupli (I - VI) (6.91 ± 0.453 days), copepodite (I - VI) (11.4 ± 0.311days) and naupli to adult (20 ± 1.08 days) appeared significantly longer when fed with Nannochloropsis sp. compared to other treatments. The daily mean naupli production of adult females over 7 days was significantly higher (p ˂ 0.05) in Nannochloropsis sp. compared with Chlamydomonas sp. and Gonyostomum sp. On the 25th day of Nannochloropsis sp. treatment, 99 % of the mature females died. Production (naupli, copepodite adult male and adult female) was significantly higher (p ˂ 0.05) in Nannochloropsis sp. than in other microalgal diets. On the fifteenth day, Nannochloropsis sp. showed a significantly higher (p ˂ 0.05) specific growth rate than other microalgal diets. Nannochloropsis sp. had the highest nauplius survival rate on the sixth day compared to other microalgal diets. With Nannochloropsis sp., the species has a higher hatching rate, and in Chlamydomonas sp. hatching occurs earlier. The average lifespan for Nannochloropsis sp. was 46 days, for Chlamydomonas sp. it was 37 days, and for Gonyostomum sp. it was 32 days.


a b s t r a c t
The effects of growth and reproduction on the marine Cyclops sp. were investigated using three microalgae as diets.The development period of Cyclops sp. was evaluated at 10 6 cells/ml in 15ppt salinity to identify the stationary phase.The survival rate of marine Cyclops from nauplius to adult differed according to the microalgal diet.The results showed that the shortest time (14 days) and highest survival (17.6 ± 0.131 %) for Cyclops sp. was achieved with those fed with Nannochloropsis sp.Whereas, it took longest time (37 days) and lowest survival rate (6.40 ± 0.035 %) when fed C hlamydomonas sp.The developmental period from naupli (I -VI) (6.91 ± 0.453 days), copepodite (I -VI) (11.4 ± 0.311days) and naupli to adult (20 ± 1.08 days) appeared significantly longer when fed with Nannochloropsis sp.compared to other treatments.The daily mean naupli production of adult females over 7 days was significantly higher ( p ˂ 0.05) in Nannochloropsis sp.compared with Chlamydomonas sp. and Gonyostomum sp.On the 25th day of Nannochloropsis sp.treatment, 99 % of the mature females died.Production (naupli, copepodite adult male and adult female) was significantly higher ( p ˂ 0.05) in Nannochloropsis sp.than in other microalgal diets.On the fifteenth day, Nannochloropsis sp.showed a significantly higher ( p ˂ 0.05) specific growth rate than other microalgal diets.
Nannochloropsis sp. had the highest nauplius survival rate on the sixth day compared to other microalgal diets.With Nannochloropsis sp ., the species has a higher hatching rate, and in Chlamydomonas sp.hatching occurs earlier.

Value of the Data
• The data will contribute to the selection of potential microalgae culture for marine Cyclops sp.• Data on population growth, developmental stages, and survival, will be useful in selecting microalgae with the best potential as live feed source in the mass production of copepod for aquaculture application.• Data on growth (population growth and survival rates) and reproductive parameters (developmental stages, sex ratio, fecundity, and egg hatchability) are important basic information in the study of copepod biology as well as guide for further investigations or applications in copepod ecology and aquaculture application.

Background
Zooplankton is a marine microorganism that forms primary and secondary linkages in the food webs of all aquatic habitats [ 2 , 3 ].Copepods are commonly used as live feed in the aquaculture sector, notably for marine fish along with crustacean larvae [ 4 ].Marine copepods are commercially cultivable species their growth and survival have been impacted by different feed-ing habit [5][6][7].Food quality and quantity are also major factors influencing copepod growth and fecundity.Copepods require algal feeding, which promotes the growth of marine Cyclops sp .The present study investigated the growth and population of marine Cyclops sp.fed with different microalgae.Although certain marine copepod species have been successfully grown in the past, there are still very few species that have been recorded.The use of Cyclops as live food has various benefits, including enhanced fish and crustacean larval survival, accelerated larval growth, and provision of essential nutrients.
There is currently no documentation on the culture of this Cyclops sp .or their parameters when given microalgal diets.The main issue with using copepods as live feed in the aquaculture sector is inconsistent production due to poor culture procedures and a lack of information about their optimal diets [ 8 ].
The total population growth rate of Cyclops sp. per dietary treatment after 25 days of culture varied based on food type ( p < 0.05, Table 2 ).Copepods fed Nannochloropsis sp.achieved highest number of naupli, copepodite and adult mane.On the other hand, Copepods fed Gonyostomum sp.grow more than Chlamydomonas sp. ( p < 0.05).Table 2 shows the population growth rate of copepods fed multiple microalgae diets.
According to the graph presented there was no significance in terms of hatching rate of female copepods by algal diet type ( p < 0.05, Fig. 2 ).Copepods fed with Nannochloropsis sp., Gonyostomum sp., Chlamydomonas sp.showed similar hatching rate (1.5 ± 0.11 %).Copepods fed Chlamydomonas sp.exhibited the development time (11.38 ± 0.139 days) compared to other dietary regimens ( p < 0.05).Copepodite maturation periods did not differ substantially between food types ( p = 0.091, Fig. 3 ).
Table 3 showed the lifespans of female marine Cyclops sp.under various dietary treatments.Copepods fed on Nannochloropsis sp. had the highest lifespan (35.2 ± 2.39 days).while those fed with Chlamydomonas sp. had the shortest at 25.0 ± 2.55 days (Table 3).
Copepods produced the most spawns in their lives when fed Nannochloropsis sp .(4.2 ± 0.836).Different diets had a significant impact on the number of spawns in a life-   3).
The number of offspring produced per egg sac per spawn varied based on diet type, where those fed with Nannochloropsis sp .achieved the highest (6.4 ± 1.14), while those fed with Chlamydomonas sp .produced the fewest among diets ( Table 3 ).

Collection of zooplankton cyclops sp . and microalgal strains
Pure strain of Nannochloropsis sp., Gonyostomum sp., and Chlamydomonas sp. and Cyclops sp. were collected from live feed research corner, Chattogram Veterinary and Animal Sciences University to conduct the experiment.

Stock culture
In stock culture, isolated species were transferred to a 1 L beaker filled with filtered seawater.During culture, 24-h aeration was provided; Cyclops sp .was fed with mixed algae at a concentration of 2 × 10 6 cells/ml for once in daily.The culture was then carried out continuously for numerous generations to ensure a mono-species stock.Additionally, wastes were siphoned and the expelled culture water was exchanged with filtered fresh seawater on daily basis.

Maintenance of microalgae culture
The purified samples were cultivated in Conway culture medium with 15 ppt salinity.The stock was scaled up and then mass-cultivated for Cyclops sp .diets.

Experimental design
The effects of microalgae diets on the reproductive development of Cyclops sp .were studied using three different microalgae Nannochloropsis sp., Gonyostomum sp., and Chlamydomonas sp. at 15 ppt salinity in the 8-hour dark and 16-h light on 24 h photoperiod.Throughout the experimental period copepod was measured and counted on daily basis under the microscope (Optika, Italy).These studies were carried out under controlled environment, with five replicates.The experiment was conducted in a petri dish with one gravid copepod and 40 ml of filtered autoclave seawater with three different microalgae diet.
The experiment starts with one adult female from the first breeding with egg sacs.They were given Nannochloropsis sp., Gonyostomum sp., and Chlamydomonas sp. with concentrations of 10 6 cells/ml maintained throughout the experiment.In the experiment, algal cells were supplied and counted daily using hemocytometer under the microscope (Optika, Italy) to maintain the same feed content.Cells were counted in the two chambers of the hemocytometer under a magnification of 40 ×.The following formula was used for the cell density calculation [ 9 ]: Cell count calculation ( cell /mL ) for 25 squares = Total number of cells counted 50 × 4 × 10 6 In this equation, 50 stood for the 50 squares within each of the two hemocytometer chambers, and 4 × 10 −6 for the volume of samples spread among the tiny square areas, which was equal to 0.004 mm 3 (0.2 mm × 0.2 mm × 0.1 mm), expressed in cm 3 (mL).
All replicates were monitored at 6-hour intervals to determine hatching and the lifespan.After the female copepods released their eggs, they were transferred to a different petri dish.A similar method was followed until the female produced more egg sacs.

Determination of population and individual growth
The experimental conditions were maintained constant for 25 days, with manual shaking every 2times per day.Each feeding treatment was replicated five times to identify the copepod population.Samples were collected from each treatment daily with replicates to calculate and record copepod development phases (naupli, copepodite, and adult) as well as copepod density.
Specific growth rate was calculated from the density data by using the following equation [ 10 ] Where, t is cultured days, Ni are the initial density of copepods and Ne is the end density of copepods.

Determination of survival rate
Survival rate of copepod was calculated by using the following equation [ 11 ]: Survival rate = ( total number of five samples taken once in two days ÷ total number of initial copepod density in petri dish ) ×100%

. Hatching time and hatching rate
The eggs were monitored at 6-hour intervals to record the newly hatched naupli over a 24-h duration.Following a period of 48 h since hatching, the unhatched egg was tallied and documented.Equation use to calculate the hatching rate is as follows [ 11 ]: Hatching rate = [ 1 −( number of unhatched eggs ÷ number of total eggs ) ]

Development time
The growth stages of copepods were observed under a microscope to determine their maturation period (from Nauplius to Copepodite, Copepodite to adult, and production of offspring from the nauplii survived from the 1st generation).

Off spring production, spawning and lifespan
After spawning, each male-female couple was transferred to a new beaker to allow for better observation of future spawning.Offspring productions were calculated using the average of triplicate pairs of cyclops sp. in each treatment.

Sex ratio
At the end of the experiment, the sex ratio was calculated by counting all male and female adults separately.Adult copepods were counted using a dissecting microscope (Nikon E600).

Quality assurance and quality control
Each instrument was calibrated by the applicable standard operating procedures (SOPs) and manufacturer instructions.Balance and other measuring equipment were regularly calibrated.Before starting the experiment, all equipment was checked properly.The tests that were undertaken produced consistent results.The experimental apparatus was cleaned with chlorine and sterilized in an autoclave before and after use.

Statistical analysis
Microsoft Excel was used to calculate the mean and standard deviation.All data were tested using a one-way analysis of variance (ANOVA).Statistical analyses were carried out using the IBM SPSS (v.26.0) software.Significant differences between groups were determined at a pvalue < 0.05.

Limitations
Not applicable.

Fig. 3 .
Fig. 3. Development time of naupli to adult with different microalgae.

4. 7 .
Determination of reproductive performance 4.7.1 The average lifespan for Nannochloropsis sp. was 46 days, for Chlamydomonas sp. it was 37 days, and for Gonyostomum sp. it was 32 days.©2024 The Author(s).Published by Elsevier Inc.
The copepod was examined at regular intervals (every 2 h) for the hatching of naupli.Once the naupli released, the adult female was carefully removed from the test tube and the naupli were counted under the microscope with Sedgewick-Rafter chamber.In this experiment, the time taken for eggs to hatch from spawning (embryonic development), the development

Table 1
Effects of Cyclops sp.survival rate (%) and specific growth rate fed with different microalgae.All values are mean ± SD ( n = 5).Different small letters indicate significant differences between treatments ( p < 0.05).

Table 2
Developmental stages (no. of individuals) and sex ratio (M : F) fed with three different microalgae.All values are mean ± SD ( n = 5).Significant differences between treatments are indicated by small letters ( p < 0.05).All of the data represents the average of individual's number except sex ratio.b Chlamydomonas sp.14.4 ± 2.97 c 10.8 ± 2.28 c 8 ± 3.74 b 2.4 ± 1.67 a 35.5 ± 15.5 b

Table 3
Effects of reproductive performance of Cyclops sp.fed with different microalgae.